Formulation for spray-drying large porous particles

ABSTRACT

Particles having a tap density less than about 0.4 g/cm3 are formed by spray drying from a colloidal solution including a carboxylic acid or salt thereof, a phospholipid, a divalent salt and a solvent such as an aqueous-organic solvent. The colloidal solution can also include a therapeutic, prophylactic or diagnostic agent. Preferred carboxylic acids include at least two carboxyl groups. Preferred phospholipids include phosphatidylcholines, phosphatidylethanolamines, phosphatidylglycerols, phophstidylserines, phosphatidylinositols and combinations thereof. The particles are suitable for pulmonary delivery.

RELATED APPLICATION

This application is a divisional of U.S. Ser. No.: 09/644,105, filedAug. 23, 2000, now U.S. Pat. No. 6,749,835 which claims the benefit ofU.S. Provisional Application No.: 60/150,662, filed on Aug. 25. 1999.

BACKGROUND OF THE INVENTION

Aerosols for the delivery of therapeutic agents to the respiratory tracthave been described, for example, Adjei, A. and Garren, J., Pharm. Res.,7: 565-569 (1990); and Zanen, P. and Lamm, J.-W. J. Int. J. Pharm., 114:111-115 (1995). The respiratory tract encompasses the upper airways,including the oropharynx and larynx, followed by the lower airways,which include the trachea followed by bifurcations into the bronchi andbronchioli. The upper and lower airways are called the conductingairways. The terminal bronchioli then divide into respiratory bronchioliwhich then lead to the ultimate respiratory zone, the alveoli, or deeplung. Gonda, I. “Aerosols for delivery of therapeutic and diagnosticagents to the respiratory tract,” in Critical Reviews in TherapeuticDrug Carrier Systems, 6: 273-313 (1990). The deep lung, or alveoli, arethe primary target of inhaled therapeutic aerosols for systemic drugdelivery.

Inhaled aerosols have been used for the treatment of local lungdisorders including asthma and cystic fibrosis (Anderson, Am. Rev.Respir. Dis., 140: 1317-1324 (1989)) and have potential for the systemicdelivery of peptides and proteins as well (Patton and Platz, AdvancedDrug Delivery Reviews, 8: 179-196 (1992)). However, pulmonary drugdelivery strategies present many difficulties for the delivery ofmacromolecules; these include protein denaturation duringaerosolization, excessive loss of inhaled drug in the oropharyngealcavity (often exceeding 80%), poor control over the site of deposition,lack of reproducibility of therapeutic results owing to variations inbreathing patterns, the frequent too-rapid absorption of drugpotentially resulting in local toxic effects, and phagocytosis by lungmacrophages.

Considerable attention has been devoted to the design of therapeuticaerosol inhalers to improve the efficiency of inhalation therapies.Timsina et. al., Int. J. Pharm., 101: 1-13 (1995); and Tansey, I. P.,Spray Technol. Market, 4: 26-29 (1994). Attention has also been given tothe design of dry powder aerosol surface texture, regarding particularlythe need to avoid particle aggregation, a phenomenon which considerablydiminishes the efficiency of inhalation therapies. French, D. L.,Edwards, D. A. and Niven, R. W., J. Aerosol Sci., 27: 769-783 (1996).Dry powder formulations (“DPFs”) with large particle size have improvedflowability characteristics, such as less aggregation (Visser, J.,Powder Technology 58: 1-10 (1989)), easier aerosolization, andpotentially less phagocytosis. Rudt, S. and R. H. Muller, J. ControlledRelease, 22: 263-272 (1992); Tabata, Y. and Y. Ikada, J. Biomed. Mater.Res., 22: 837-858 (1988). Dry powder aerosols for inhalation therapy aregenerally produced with mean geometric diameters primarily in the rangeof less than 5 μm. Ganderton, D., J. Biopharmaceutical Sciences, 3:101-105 (1992); and Gonda, I. “Physico-Chemical Principles in AerosolDelivery,” in Topics in Pharmaceutical Sciences 1991, Crommelin, D. J.and Midha, K. K., Eds., Medpharm Scientific Publishers, Stuttgart, pp.95-115, 1992. Large “carrier” particles (containing no drug) have beenco-delivered with therapeutic aerosols to aid in achieving efficientaerosolization among other possible benefits. French, D. L., Edwards, D.A. and Niven, R. W., J. Aerosol Sci., 27: 769-783 (1996).

The human lungs can remove or rapidly degrade hydrolytically cleavabledeposited aerosols over periods ranging from minutes to hours. In theupper airways, ciliated epithelia contribute to the “mucociliaryescalator” by which particles are swept from the airways toward themouth. Pavia, D. “Lung Mucociliary Clearance,” in Aerosols and the Lung:Clinical and Experimental Aspects, Clarke, S. W. and Pavia, D., Eds.,Butterworths, London, 1984. Anderson, Am. Rev. Respir. Dis., 140:1317-1324 (1989). In the deep lungs, alveolar macrophages are capable ofphagocytosing particles soon after their deposition. Warheit, M. B. andHartsky, M. A., Microscopy Res. Tech., 26: 412-422 (1993); Brain, J. D.,“Physiology and Pathophysiology of Pulmonary Macrophages,” in TheReticuloendothelial System, Reichard, S. M. and Filkins, J., Eds.,Plenum, New York, pp. 315-327, 1985; Dorries, A. M. and Valberg, P. A.,Am. Rev. Resp. Disease 146: 831-837 (1991); and Gehr, P., MicroscopyRes. and Tech., 26: 423-436 (1993). As the diameter of particles exceeds3 μm, there is increasingly less phagocytosis by macrophages. Kawaguchi,H., Biomaterials 7: 61-66 (1986); Krenis, L. J. and Strauss, B., Proc.Soc. Exp. Med., 107: 748-750 (1961); and Rudt, S. and Muller, R. H., J.Contr. Rel., 22: 263-272 (1992). However, increasing the particle sizealso has been found to minimize the probability of particles (possessingstandard mass density) entering the airways and acini due to excessivedeposition in the oropharyngeal or nasal regions. Heyder, J., J. AerosolSci., 17: 811-825 (1986).

Local and systemic inhalation therapies can often benefit from arelatively slow controlled release of the therapeutic agent. Gonda, I.,“Physico-chemical principles in aerosol delivery,” in: Topics inPharmaceutical Sciences 1991, D. J. A. Crommelin and K. K. Midha, Eds.,Stuttgart: Medpharm Scientific Publishers, pp. 95-117 (1992). Slowrelease from a therapeutic aerosol can prolong the residence of anadministered drug in the airways or acini, and diminish the rate of drugappearance in the bloodstream. Also, patient compliance is increased byreducing the frequency of dosing. Langer, R., Science, 249: 1527-1533(1990); and Gonda, I., “Aerosols for delivery of therapeutic anddiagnostic agents to the respiratory tract,” in Critical Reviews inTherapeutic Drug Carrier Systems 6: 273-313 (1990).

Controlled release drug delivery to the lung may simplify the way inwhich many drugs are taken. Gonda, I., Adv. Drug Del. Rev., 5: 1-9(1990); and Zeng, X., et al., Int. J. Pharm., 124: 149-164 (1995).Pulmonary drug delivery is an attractive alternative to oral,transdermal, and parenteral administration because self-administrationis simple, the lungs provide a large mucosal surface for drugabsorption, there is no first-pass liver effect of absorbed drugs, andthere is reduced enzymatic activity and pH mediated drug degradationcompared with the oral route. Relatively high bioavailability of manymolecules, including macromolecules, can be achieved via inhalation.Wall, D. A., Drug Delivery, 2: 1-20 1995); Patton, J. and Platz, R.,Adv. Drug Del. Rev., 8: 179-196 (1992); and Byron, P., Adv. Drug. Del.Rev., 5: 107-132 (1990). As a result, several aerosol formulations oftherapeutic drugs are in use or are being tested for delivery to thelung. Patton, J. S., et al., J. Controlled Release, 28: 79-85 (1994);Damms, B. and Bains, W., Nature Biotechnology (1996); Niven, R. W., etal., Pharm. Res., 12(9): 1343-1349 (1995); and Kobayashi, S., et al.,Pharm. Res., 13(1): 80-83 (1996).

Drugs currently administered by inhalation come primarily as liquidaerosol formulations. However, many drugs and excipients, especiallyproteins, peptides (Liu, R., et al., Biotechnol. Bioeng., 37: 177-184(1991)), and biodegradable carriers such as poly(lactide-co-glycolides)(PLGA), are unstable in aqueous environments for extended periods oftime. This can make storage as a liquid formulation problematic. Inaddition, protein denaturation can occur during aerosolization withliquid formulations. Mumenthaler, M., et al., Pharm. Res., 11: 12-20(1994). Considering these and other limitations, dry powder formulations(DPF's) are gaining increased interest as aerosol formulations forpulmonary delivery. Damms, B. and Bains, W., Nature Biotechnology(1996); Kobayashi, S., et al., Pharm. Res., 13(1): 80-83 (1996); andTimsina, M., et al., Int. J. Pharm., 101: 1-13 (1994). However, amongthe disadvantages of DPF's is that powders of ultrafine particulatesusually have poor flowability and aerosolization properties, leading torelatively low respirable fractions of aerosol, which are the fractionsof inhaled aerosol that escape deposition in the mouth and throat.Gonda, I., in Topics in Pharmaceutical Sciences 1991, D. Crommelin andK. Midha, Editors, Stuttgart: Medpharm Scientific Publishers, 95-117(1992). A primary concern with many aerosols is particulate aggregationcaused by particle-particle interactions, such as hydrophobic,electrostatic, and capillary interactions. An effective dry-powderinhalation therapy for both short and long term release of therapeutics,either for local or systemic delivery, requires a powder that displaysminimum aggregation, as well as a means of avoiding or suspending thelung's natural clearance mechanisms until drugs have been effectivelydelivered.

Therefore, a need exists for dry-powders suitable for inhalation whichminimize or eliminate the above-mentioned problems.

SUMMARY OF THE INVENTION

The invention relates to particles having a tap density of less thanabout 0.4 g/cm³ and preferably less than about 0.1 g/cm³. The particlesinclude a carboxylate group or moiety. The particles further include amultivalent salt or its ionic components. In one embodiment of theinvention, the particles further include a phospholipid. In addition,the particles can include a therapeutic, prophylactic or diagnosticagent or any combination thereof. In one embodiment, the particles havea median geometric diameter of between about 5 microns (μm) and about 30μm, preferably at least about 9 μm. In another embodiment, the particleshave an aerodynamic diameter of between about 1 μm and about 5 μm.

The invention also relates to a method of producing particles having atap density of less than about 0.4 g/cm³. The method includes forming amixture which includes a carboxylate moiety, such as provided, forexample, by a carboxylic acid or salt thereof, a multivalent salt, aphospholipid, and a solvent. The mixture can also include a therapeutic,prophylactic or diagnostic agent, or any combination thereof. Themixture is spray-dried to form particles having a tap density of lessthan about 0.4 g/cm³. Preferred solvents that can be employed in thespray drying process include organic or organic-aqueous solvents. In apreferred embodiment, the mixture fed to the spray drying apparatus is acolloidal suspension.

The invention further relates to a method of delivering a therapeutic,prophylactic or diagnostic agent to the pulmonary system of a patient inneed of treatment, prophylaxis or diagnosis. The method includesadministering to the respiratory tract of the patient an effectiveamount of particles having a tap density of less than about 0.4 g/cm³and preferably less than about 0.1 g/cm³. The particles include atherapeutic, prophylactic or diagnostic agent, or any combinationthereof and a carboxylate moiety. The particles further include amultivalent salt or its ionic components. In one embodiment of theinvention, the particles also include a phospholipid. Delivery to therespiratory system can be primarily to the deep lung, to the centralairways or to the upper airways.

The invention relates also to a composition for delivery to a patient inneed of treatment, prophylaxis or diagnosis. The composition includesparticles which have a tap density of less than about 0.4 g/cm³ andpreferably less than about 0.1 g/cm³. In one embodiment, the particlesinclude a carboxylate moiety, a multivalent salt and a phospholipid. Ina preferred embodiment, the particles also include a therapeutic,prophylactic or diagnostic agent. In another preferred embodiment,delivery is to the pulmonary system.

In a preferred embodiment, the carboxylate moiety is a hydrophiliccarboxylic acid or salt thereof. In another embodiment, preferredcarboxylate moieties include at least two carboxyl groups.

In a preferred embodiment, the salt is a divalent salt. Suitabledivalent salts include, for example chlorides of alkaline earth metals.Calcium chloride (CaCl₂) is preferred. In another preferred embodiment,the multivalent salt is a pharmaceutically acceptable salt.

Preferred phospholipids include but are not limited to phosphatidicacid, phosphatidylcholines, phosphatidylethanolamines,phosphatidylglycerols, phosphatidylserines, phosphatidylinositols andcombinations thereof.

The invention has several advantages. Pulmonary delivery advantageouslycan reduce or eliminate the need for injection. For example, therequirement for daily insulin injections can be avoided. Furthermore,the particles of the invention can be delivered as a dry powder to thedeep lung, upper or central airways. They can be used to providecontrolled systemic or local delivery of therapeutic or diagnosticagents to the respiratory tract via aerosolization. The particles can beeasily prepared from simple, lung-compatible compounds without requiringthe use of large macromolecules such as polymers, proteins,polysaccharides and others. The formation of colloidal suspensionsresults in particles of desired shape and porosity. Compared to methodsthat require solubilizing, higher concentrations can be employed.Administration of the particles to the lung by aerosolization permitsdeep lung delivery of relatively large diameter therapeutic aerosols,for example, greater than about 5 μm in mean diameter. The particles canbe fabricated with a rough surface texture to reduce particleagglomeration and improve flowability of the powder. The spray-driedparticle can be fabricated with features which enhance aerosolizationvia dry powder inhaler devices, and lead to lower deposition in themouth, throat and inhaler device.

DETAILED DESCRIPTION OF THE INVENTION

The features and other details of the invention, either as steps of theinvention or as combination of parts of the invention, will now be moreparticularly described and pointed out in the claims. It will beunderstood that the particular embodiments of the invention are shown byway of illustration and not as limitations of the invention. Theprinciple feature of this invention may be employed in variousembodiments without departing from the scope of the invention.

The invention is directed to particles having a tap density of less thanabout 0.4 g/cm³ and preferably less than about 0.1 g/cm³ and to methodsof producing such particles. The particles can be employed for deliveryof a therapeutic, prophylactic or diagnostic agent to a patient in needof therapy, prophylaxis or diagnosis. In a preferred embodiment,delivery is to the pulmonary system. The particles can also be deliveredto nonhuman mammals such as, for example, to laboratory animals or inveterinary medicine.

The particles include a carboxylate moiety. In one embodiment of theinvention, the carboxylate moiety includes at least two carboxyl groups.Carboxylate moieties can be provided by carboxylic acids, salts thereofas well as by combinations of two or more carboxylic acids and/or saltsthereof. In a preferred embodiment, the carboxylate moiety is ahydrophilic carboxylic acid or salt thereof. Suitable carboxylic acidsinclude but are not limited to hydroxydicarboxylic acids,hydroxytricarboxilic acids and the like. Citric acid and citrates, suchas, for example sodium citrate, are preferred. Combinations or mixturesof carboxylic acids and/or their salts also can be employed.

The carboxylate moiety can be present in the particles in an amountranging from about 10 to about 80% weight. Preferably, the carboxylatemoiety can be present in the particles in an amount 10-20%.

The particles also include a multivalent salt or its ionic components.As used herein, a “multivalent” salt includes divalent salts. In apreferred embodiment, the salt is a divalent salt. In another preferredembodiment, the salt is a salt of an alkaline-earth metal, such as, forexample, calcium chloride. The particles of the invention can alsoinclude mixtures or combinations of salts and/or their ionic components.

The salt or its ionic components are present in the particles in anamount ranging from about 5 to about 40% weight.

The particles further include a phospholipid, also referred to herein asphosphoglyceride. In a preferred embodiment, the phospholipid, isendogenous to the lung. In another preferred embodiment the phospholipidincludes, among others, phosphatidic acid, phosphatidylcholines,phosphatidylethanolamines, phosphatidylglycerols, phophatidylserines,phosphatidylinositols and combinations thereof. Specific examples ofphospholipids include but are not limited to phosphatidylcholinesdipalmitoyl phosphatidylcholine (DPPC), dipalmitoylphosphatidylethanolamine (DPPE), distearoyl phosphatidylcholine (DSPC),dipalmitoyl phosphatidyl glycerol (DPPG) or any combination thereof.

The phospholipid can be present in the particles in an amount rangingfrom about 20 to about 90% weight. Preferably, it can be present in theparticles in an amount ranging from about 50 to about 80% weight.

Suitable methods of preparing and administering particles which includephospholipids, are described in U.S. Pat. No. 5,855,913, issued on Jan.5, 1999 to Hanes et al. and in U.S. Pat. No. 5,985,309, issued on Nov.16, 1999 to Edwards et al. The teachings of both are incorporated hereinby reference in their entirety.

In another embodiment of the invention the particles include asurfactant such as, but not limited to the phospholipids describedabove. Other surfactants, such as, for example, hexadecanol; fattyalcohols such as polyethylene glycol (PEG); polyoxyethylene-9-laurylether; a surface active fatty acid, such as palmitic acid or oleic acid;glycocholate; surfactin; a poloxomer; a sorbitan fatty acid ester suchas sorbitan trioleate (Span 85); tyloxapol can also be employed.

As used herein, the term “surfactant” refers to any agent whichpreferentially absorbs to an interface between two immiscible phases,such as the interface between water and an organic polymer solution, awater/air interface or organic solvent/air interface. Surfactantsgenerally possess a hydrophilic moiety and a lipophilic moiety, suchthat, upon absorbing to microparticles, they tend to present moieties tothe external environment that do not attract similarly-coated particles,thus reducing particle agglomeration. Surfactants may also promoteabsorption of a therapeutic or diagnostic agent and increasebioavailability of the agent.

The surfactant can be present in the particles in an amount ranging fromabout 20 to about 90. Preferably, it can be present in the particles inan amount ranging from about 50 to about 80.

Examples of therapeutic, prophylactic or diagnostic agents, alsoreferred to herein as “bioactive agents”, “drugs” or “medicaments”, bothlocally as well as systemically acting agents. The particles can alsoinclude mixtures of therapeutic, prophylactic and/or diagnostic agents.Furthermore, the particles can include needed biological compounds suchas, for example, blood, plasma or oxygen. The particles can includehydrophilic as well as hydrophobic drugs.

Examples of therapeutic, prophylactic or diagnostic agents include, butare not limited to synthetic inorganic and organic compounds, proteins,peptides, polypeptides, polysaccharides and other sugars, lipids, andDNA and RNA nucleic acid sequences having therapeutic, prophylactic ordiagnostic activities. Nucleic acid sequences include genes, antisensemolecules which bind to complementary DNA or RNA and inhibittranscription, and ribozymes. Polysaccharides, such as heparin, can alsobe administered. The agents to be incorporated can have a variety ofbiological activities, such as vasoactive agents, neuroactive agents,hormones, anticoagulants, immunomodulating agents, cytotoxic agents,prophylactic agents, antibiotics, antivirals, antisense, antigens, andantibodies. In some instances, the proteins may be antibodies orantigens which otherwise would have to be administered by injection toelicit an appropriate response. Compounds with a wide range of molecularweight can be encapsulated, for example, between 100 and 500,000 gramsor more per mole.

Proteins are defined as consisting of 100 amino acid residues or more;peptides are less than 100 amino acid residues. Unless otherwise stated,the term protein refers to both proteins and peptides. Examples includeinsulin and other hormones.

Those therapeutic agents which are charged, such as most of theproteins, including insulin, can be administered as a complex betweenthe charged therapeutic agent and a molecule of opposite charge.Preferably, the molecule of opposite charge is a charged lipid or anoppositely charged protein.

The particles can include a therapeutic agent for local delivery withinthe lung, such as agents for the treatment of asthma, chronicobstructive pulmonary disease (COPD), emphysema, or cystic fibrosis, orfor systemic treatment. For example, genes for the treatment of diseasessuch as cystic fibrosis can be administered, as can beta agonists,steroids, anticholinergies, and leukotriene modifers for asthma. Otherspecific therapeutic agents include, but are not limited to, humangrowth hormone, insulin, calcitonin, gonadotropin-releasing hormone(“LHRH”), granulocyte colony-stimulating factor (“G-CSF”), parathyroidhormone-related peptide, somatostatin, testosterone, progesterone,estradiol, nicotine, fentanyl, norethisterone, clonidine, scopolamine,salicylate, cromolyn sodium, salmeterol, formeterol, albuterol, andValium.

The particles can include any of a variety of diagnostic agents tolocally or systemically deliver the agents following administration to apatient.

Diagnostic agents also include but are not limited to imaging agentswhich include commercially available agents used in positron emissiontomography (PET), computer assisted tomography (CAT), single photonemission computerized tomography, x-ray, fluoroscopy, and magneticresonance imaging (MRI).

Examples of suitable materials for use as contrast agents in MRI includebut are not limited to the gadolinium chelates currently available, suchas diethylene triamine pentacetic acid (DTPA) and gadopentotatedimeglumine, as well as iron, magnesium, manganese, copper and chromium.

Examples of materials useful for CAT and x-rays include iodine basedmaterials for intravenous administration, such as ionic monomerstypified by diatrizoate and iothalamate, non-ionic monomers such asiopamidol, isohexol, and ioversol, non-ionic dimers, such as iotrol andiodixanol, and ionic dimers, for example, ioxagalte.

Preferably, a therapeutic agent can be present in the spray-driedparticles in an amount ranging from less than about 1% to about 40%.Preferably, a prophylactic agent can be present in the spray-driedparticles in an amount ranging from about less than about 1% to about40%. Preferably, a diagnostic agent can be present in the spray-driedparticles in an amount ranging from about less than about 1% to about40%.

In one embodiment of the invention, the phospholipid or combination orphospholipids present in the particles can have a therapeutic,prophylactic or diagnostic role. For example, the particles of theinvention can be used to deliver surfactants to the lung of a patient.This is particularly useful in medical indications which requiresupplementing or replacing endogenous lung surfactants, for example inthe case of infant respiratory distress syndrome.

The particles can include other materials. In one embodiment of theinvention, the particles also include an amino acid. Hydrophobic aminoacids are preferred. Suitable amino acids include naturally occurringand non-naturally occurring hydrophobic amino acids. Some suitablenaturally occurring hydrophobic amino acids, include but are not limitedto, leucine, isoleucine, alanine, valine, phenylalanine, glycine andtryptophan. Combinations of hydrophobic amino acids can also beemployed. Non-naturally occurring amino acids include, for example,beta-amino acids. Both D and L configurations and racemic mixtures ofhydrophobic amino acids can be employed. Suitable hydrophobic aminoacids can also include amino acid derivatives or analogs. As usedherein, an amino acid analog includes the D or L configuration of anamino acid having the following formula: —NH—CHR—CO—, wherein R is analiphatic group, a substituted aliphatic group, a benzyl group, asubstituted benzyl group, an aromatic group or a substituted aromaticgroup and wherein R does not correspond to the side chain of anaturally-occurring amino acid. As used herein, aliphatic groups includestraight chained, branched or cyclic C1-C8 hydrocarbons which arecompletely saturated, which contain one or two heteroatoms such asnitrogen, oxygen or sulfur and/or which contain one or more units ofunsaturation. Aromatic groups include carbocyclic aromatic groups suchas phenyl and naphthyl and heterocyclic aromatic groups such asimidazolyl, indolyl, thienyl, furanyl, pyridyl, pyranyl, oxazolyl,benzothienyl, benzofuranyl, quinolinyl, isoquinolinyl and acridintyl.

Suitable substituents on an aliphatic, aromatic or benzyl group include—OH, halogen (—Br, —Cl, —I and —F) —O(aliphatic, substituted aliphatic,benzyl, substituted benzyl, aryl or substituted aryl group), —CN, —NO₂,—COOH, —NH₂, —NH(aliphatic group, substituted aliphatic, benzyl,substituted benzyl, aryl or substituted aryl group), —N(aliphatic group,substituted aliphatic, benzyl, substituted benzyl, aryl or substitutedaryl group)₂, —COO(aliphatic group, substituted aliphatic, benzyl,substituted benzyl, aryl or substituted aryl group), —CONH₂,—CONH(aliphatic, substituted aliphatic group, benzyl, substitutedbenzyl, aryl or substituted aryl group)), —SH, —S(aliphatic, substitutedaliphatic, benzyl, substituted benzyl, aromatic or substituted aromaticgroup) and —NH—C(═NH)—NH₂. A substituted benzylic or aromatic group canalso have an aliphatic or substituted aliphatic group as a substituent.A substituted aliphatic group can also have a benzyl, substitutedbenzyl, aryl or substituted aryl group as a substituent. A substitutedaliphatic, substituted aromatic or substituted benzyl group can have oneor more substituents. Modifying an amino acid substituent can increase,for example, the lypophilicity or hydrophobicity of natural amino acidswhich are hydrophilic.

A number of the suitable amino acids, amino acids analogs and saltsthereof can be obtained commercially. Others can be synthesized bymethods known in the art. Synthetic techniques are described, forexample, in Green and Wuts, “Protecting Groups in Organic Synthesis”,John Wiley and Sons, Chapters 5 and 7, 1991.

Hydrophobicity is generally defined with respect to the partition of anamino acid between a nonpolar solvent and water. Hydrophobic amino acidsare those acids which show a preference for the nonpolar solvent.Relative hydrophobicity of amino acids can be expressed on ahydrophobicity scale on which glycine has the value 0.5. On such ascale, amino acids which have a preference for water have values below0.5 and those that have a preference for nonpolar solvents have a valueabove 0.5. As used herein, the term hydrophobic amino acid refers to anamino acid that, on the hydrophobicity scale has a value greater orequal to 0.5, in other words, has a tendency to partition in thenonpolar acid which is at least equal to that of glycine.

Examples of amino acids which can be employed include, but are notlimited to: glycine, proline, alanine, cysteine, methionine, valine,leucine, tyrosine, isoleucine, phenylalanine, tryptophan. Preferredhydrophobic amino acids include leucine, isoleucine, alanine, valine,phenylalanine, glycine and tryptophan. Combinations of hydrophobic aminoacids can also be employed. Furthermore, combinations of hydrophobic andhydrophilic (preferentially partitioning in water) amino acids, wherethe overall combination is hydrophobic, can also be employed.Combinations of one or more amino acids and one or more phospholipids orsurfactants can also be employed. Materials which impart fast releasekinetics to the medicament are preferred.

The amino acid can be present in the particles of the invention in anamount of about 60 weight %. Preferably, the amino acid can be presentin the particles in an amount ranging from about 5 to about 30 weight %.The salt of a hydrophobic amino acid can be present in the particles ofthe invention in an amount of about 60 weight %. Preferably, the aminoacid salt is present in the particles in an amount ranging from about 5to about 30 weight %. Methods of forming and delivering particles whichinclude an amino acid are described in U.S. patent application Ser. No.09/382,959, filed on Aug. 25, 1999, entitled Use of Simple Amino Acidsto Form Porous Particles During Spray Drying, and U.S. patentapplication Ser. No. 09/644,320 filed concurrently herewith and entitledUse of Simple Amino Acids to Form Porous Particles; the teachings ofboth are incorporated herein by reference in their entirety.

The particles of the invention can have desired drug release properties.In one embodiment, the particles include one or more phospholipidsselected according to their transition temperature. For example, byadministering particles which include a Phospholipid or combination ofphospholipids which have a phase transition temperature higher than thepatient's body temperature, the release of the therapeutic, prophylacticor diagnostic agent can be slowed down. On the other hand, rapid releasecan be obtained by including in the particles phospholipids having lowtransition temperatures. Particles having controlled release propertiesand methods of modulating release of a biologically active agent aredescribed in U.S Provisional Application No. 60/150,742, entitledModulation of Release From Dry Powder Formulations by Controlling MatrixTransition, filed on Aug. 25, 1999 and U.S. patent application Ser. No.10/425,194 filed concurrently herewith under entitled Modulation ofRelease From Dry Powder Formulations ; the contents of both areincorporated herein by reference in their entirety.

Particles, and in particular particles having controlled or sustainedrelease properties, also can include other materials. For example, thespray-dried particles can include a biocompatible, and preferablybiodegradable polymer, copolymer, or blend. Such polymers are described,for example, in U.S. Pat. No. 5,874,064, issued on Feb. 23, 1999 toEdwards et al., the teachings of which are incorporated herein byreference in their entirety. Preferred polymers are those which arecapable of forming aerodynamically light particles having a tap densityless than about 0.4 g/cm³, a mean diameter between about 5 μm and about30 μm and an aerodynamic diameter between approximately one and fivemicrons, preferably between one and three microns. The polymers can betailored to optimize different characteristics of the particleincluding: i) interactions between the agent to be delivered and thepolymer to provide stabilization of the agent and retention of activityupon delivery; ii) rate of polymer degradation and, thereby, rate ofdrug release profiles; iii) surface characteristics and targetingcapabilities via chemical modification; and iv) particle porosity.

Surface eroding polymers such as polyanhydrides can be used to form theparticles. For example, polyanhydrides such aspoly[(p-carboxyphenoxy)-hexane anhydride] (PCPH) may be used. Suitablebiodegradable polyanhydrides are described in U.S. Pat. No. 4,857,311.

In another embodiment, bulk eroding polymers such as those based onpolyesters including poly(hydroxy acids) can be used. For example,polyglycolic acid (PGA), polylactic acid (PLA), or copolymers thereofmay be used to form the particles. The polyester may also have a chargedor functionalizable group, such as an amino acid. In a preferredembodiment, particles with controlled release properties can be formedof poly(D,L-lactic acid) and/or poly(D,L-lactic-co-glycolic acid)(“PLGA”) which incorporate a surfactant such as DPPC.

Still other polymers include but are not limited to polyamides,polycarbonates, polyalkylenes such as polyethylene, polypropylene,poly(ethylene glycol), poly(ethylene oxide), poly(ethyleneterephthalate), poly vinyl compounds such as polyvinyl alcohols,polyvinyl ethers, and polyvinyl esters, polymers of acrylic andmethacrylic acids, celluloses and other polysaccharides, and peptides,or proteins, or copolymers or blends thereof. Polymers may be selectedwith or modified to have the appropriate stability and degradation ratesin vivo for different controlled drug delivery applications.

In one embodiment, the particles include functionalized polyester graftcopolymers, as described in Hrkach et al., Macromolecules, 28: 4736-4739(1995); and Hrkach et al., “Poly(L-Lactic acid-co-amino acid) GraftCopolymers: A Class of Functional Biodegradable Biomaterials” inHydrogels and Biodegradable Polymers for Bioapplications, ACS SymposiumSeries No. 627, Raphael M. Ottenbrite et al., Eds., American ChemicalSociety, Chapter 8, pp. 93-101, 1996.

The particles can also include other materials such as, for example,buffer salts, dextran, polysaccharides, lactose, trehalose,cyclodextrins, proteins, peptides, polypeptides, fatty acids, inorganiccompounds, phosphates.

In a preferred embodiment, the particles of the invention have a tapdensity less than about 0.4 g/cm³. Particles which have a tap density ofless than about 0.4 g/cm³ are referred herein as “aerodynamically lightparticles”. More preferred are particles having a tap density less thanabout 0.1 g/cm³. Tap density can be measured by using instruments knownto those skilled in the art such as the Dual Platform MicroprocessorControlled Tap Density Tester (Vankel, N.C.) or a GeoPyc™ instrument(Micrometrics Instrument Corp., Norcross, Ga. 30093). Tap density is astandard measure of the envelope mass density. Tap density can bedetermined using the method of USP Bulk Density and Tapped Density,United States Pharmacopia convention, Rockville, Md., 10^(th)Supplement, 4950-4951, 1999. Features which can contribute to low tapdensity include irregular surface texture and porous structure.

The envelope mass density of an isotropic particle is defined as themass of the particle divided by the minimum sphere envelope volumewithin which it can be enclosed. In one embodiment of the invention, theparticles have an envelope mass density of less than about 0.4 g/cm³.

Aerodynamically light particles have a preferred size, e.g., a volumemedian geometric diameter (VMGD) of at least about 5 microns (μm). Inone embodiment, the VMGD is from about 5 μm to about 30 μm. In anotherembodiment of the invention, the particles have a VMGD of at least 9 μm.In other embodiments, the particles have a median diameter, mass mediandiameter (MMD), a mass median envelope diameter (MMED) or a mass mediangeometric diameter (MMGD) of at least 5 μm, for example from about 5 μmand about 30 μm.

The diameter of the particles, for example, their MMGD or their VMGD,can be measured using an electrical zone sensing instrument such as aMultisizer IIe, (Coulter Electronic, Luton, Beds, England), or a laserdiffraction instrument (for example Helos, manufactured by Sympatec,Princeton, N.J.). Other instruments for measuring particle diameter arewell known in the art. The diameter of particles in a sample will rangedepending upon factors such as particle composition and methods ofsynthesis. The distribution of size of particles in a sample can beselected to permit optimal deposition within targeted sites within therespiratory tract.

Aerodynamically light particles preferably have “mass median aerodynamicdiameter” (MMAD), also referred to herein as “aerodynamic diameter”,between about 1 μm and about 5 μm. In one embodiment of the invention,the MMAD is between about 1 μm and about 3 μm. In another embodiment,the MMAD is between about 3 μm and about 5 μm.

Experimentally, aerodynamic diameter can be determined by employing agravitational settling method, whereby the time for an ensemble ofparticles to settle a certain distance is used to infer directly theaerodynamic diameter of the particles. An indirect method for measuringthe mass median aerodynamic diameter (MMAD) is the multi-stage liquidimpinger (MSLI).

The aerodynamic diameter, d_(aer), can be calculated from the equation:d _(aer) =d _(g)√ρ tapwhere d_(g) is the geometric diameter, for example the MMGD and ρ is thepowder density.

Particles which have a tap density less than about 0.4 g/cm³, mediandiameters of at least about 5 μm, and an aerodynamic diameter of betweenabout 1 μm and about 5 μm, preferably between about 1 μm and about 3 μm,are more capable of escaping inertial and gravitational deposition inthe oropharyngeal region, and are targeted to the airways or the deeplung. The use of larger, more porous particles is advantageous sincethey are able to aerosolize more efficiently than smaller, denseraerosol particles such as those currently used for inhalation therapies.

In comparison to smaller particles the larger aerodynamically lightparticles, preferably having a VMGD of at least about 5 μm, also canpotentially more successfully avoid phagocytic engulfment by alveolarmacrophages and clearance from the lungs, due to size exclusion of theparticles from the phagocytes' cytosolic space. Phagocytosis ofparticles by alveolar macrophages diminishes precipitously as particlediameter increases beyond about 3 μm. Kawaguchi, H., et al.,Biomaterials 7: 61-66 (1986); Krenis, L. J. and Strauss, B., Proc. Soc.Exp. Med., 107: 748-750 (1961); and Rudt, S. and Muller, R. H., J.Contr. Rel., 22: 263-272 (1992). For particles of statisticallyisotropic shape, such as spheres with rough surfaces, the particleenvelope volume is approximately equivalent to the volume of cytosolicspace required within a macrophage for complete particle phagocytosis.

The particles may be fabricated with the appropriate material, surfaceroughness, diameter and tap density for localized delivery to selectedregions of the respiratory tract such as the deep lung or upper orcentral airways. For example, higher density or larger particles may beused for upper airway delivery, or a mixture of varying sized particlesin a sample, provided with the same or different therapeutic agent maybe administered to target different regions of the lung in oneadministration. Particles having an aerodynamic diameter ranging fromabout 3 to about 5 μm are preferred for delivery to the central andupper airways. Particles having an aerodynamic diameter ranging fromabout 1 to about 3 μm are preferred for delivery to the deep lung.

Inertial impaction and gravitational settling of aerosols arepredominant deposition mechanisms in the airways and acini of the lungsduring normal breathing conditions. Edwards, D. A., J. Aerosol Sci., 26:293-317 (1995). The importance of both deposition mechanisms increasesin proportion to the mass of aerosols and not to particle (or envelope)volume. Since the site of aerosol deposition in the lungs is determinedby the mass of the aerosol (at least for particles of mean aerodynamicdiameter greater than approximately 1 μm), diminishing the tap densityby increasing particle surface irregularities and particle porositypermits the delivery of larger particle envelope volumes into the lungs,all other physical parameters being equal.

The low tap density particles have a small aerodynamic diameter incomparison to the actual envelope sphere diameter. The aerodynamicdiameter, d_(aer), is related to the envelope sphere diameter, d (Gonda,I., “Physico-chemical principles in aerosol delivery,” in Topics inPharmaceutical Sciences 1991 (eds. D. J. A. Crommelin and K. K. Midha),pp. 95-117, Stuttgart: Medpharm Scientific Publishers, 1992)), by theformula:d _(aer) =d√ρwhere the envelope mass p is in units of g/cm³. Maximal deposition ofmonodispersed aerosol particles in the alveolar region of the human lung(˜60%) occurs for an aerodynamic diameter of approximately d_(aer)=3 μm.Heyder, J. et al., J. Aerosol Sci., 17: 811-825 (1986). Due to theirsmall envelope mass density, the actual diameter d of aerodynamicallylight particles comprising a monodisperse inhaled powder that willexhibit maximum deep-lung deposition is:d=3/√ρμm (where ρ<1 g/cm³);where d is always greater than 3 μm. For example, aerodynamically lightparticles that display an envelope mass density, ρ=0.1 g/cm³, willexhibit a maximum deposition for particles having envelope diameters aslarge as 9.5 μm. The increased particle size diminishes interparticleadhesion forces. Visser, J., Powder Technology, 58: 1-10. Thus, largeparticle size increases efficiency of aerosolization to the deep lungfor particles of low envelope mass density, in addition to contributingto lower phagocytic losses.

The aerodyanamic diameter can be calculated to provide for maximumdeposition within the lungs, previously achieved by the use of verysmall particles of less than about five microns in diameter, preferablybetween about one and about three microns, which are then subject tophagocytosis. Selection of particles which have a larger diameter, butwhich are sufficiently light (hence the characterization“aerodynamically light”), results in an equivalent delivery to thelungs, but the larger size particles are not phagocytosed. Improveddelivery can be obtained by using particles with a rough or unevensurface relative to those with a smooth surface.

In another embodiment of the invention, the particles have an envelopemass density, also referred to herein as “mass density” of less thanabout 0.4 g/cm³. Particles also having a mean diameter of between about5 μm and about 30 μm are preferred. Mass density and the relationshipbetween mass density, mean diameter and aerodynamic diameter arediscussed in U.S. application Ser. No. 08/655,570, filed on May 24,1996, which is incorporated herein by reference in its entirety. In apreferred embodiment, the aerodynamic diameter of particles having amass density less than about 0.4 g/cm³ and a mean diameter of betweenabout 5 μm and about 30 μm is between about 1 μm and about 5 μm.

Suitable particles can be fabricated or separated, for example byfiltration or centrifugation, to provide a particle sample with apreselected size distribution. For example, greater than about 30%, 50%,70%, or 80% of the particles in a sample can have a diameter within aselected range of at least about 5 μm. The selected range within which acertain percentage of the particles must fall may be for example,between about 5 and about 30 μm, or optimally between about 5 and about15 μm. In one preferred embodiment, at least a portion of the particleshave a diameter between about 9 and about 11 μm. Optionally, theparticle sample also can be fabricated wherein at least about 90%, oroptionally about 95% or about 99%, have a diameter within the selectedrange. The presence of the higher proportion of the aerodynamicallylight, larger diameter particles in the particle sample enhances thedelivery of therapeutic or diagnostic agents incorporated therein to thedeep lung. Large diameter particles generally mean particles having amedian geometric diameter of at least about 5 μm.

The invention also relates to methods of preparing particles having atap density less than about 0.4 g/cm³. In one embodiment, the methodincludes spray drying a mixture, also referred to herein as a “feedsolution”, “feed suspension”, or “feed colloidal suspension” whichincludes a carboxylic acid or salt thereof, a phospholipid orcombination of phospholipids, a multivalent salt and a solvent. In oneembodiment, the mixture also includes a therapeutic, prophylactic ordiagnostic agent.

Suitable carboxylic acids or salts thereof include, but are not limitedto those described above. The amount of carboxylic acid or salt thereofpresent in the mixture ranges from about 10 to about 80% weight. Thephospholipid or mixture of phospholipids includes, for example, thephospholipids described above. The amount of phospholipid present in themixture ranges from about 20 to about 90% weight. The multivalent saltincludes but is not limited to the multivalent salts described above.The amount of multivalent salt present in the mixture ranges from about5 to about 40% weight. The therapeutic, prophylactic or diagnostic agentincludes but is not limited to the therapeutic, prophylactic ordiagnostic agents described above. The amount of therapeutic,prophylactic or diagnostic agent present in the mixture ranges fromabout less than 1% to about 40% weight.

Suitable organic solvents that can be employed include but are notlimited to alcohols such as, for example, ethanol, methanol, propanol,isopropanol, butanols, and others. Other organic solvents include butare not limited to perfluorocarbons, dichloromethane, chloroform, ether,ethyl acetate, methyl tert-butyl ether and others. Co-solvents that canbe employed include an aqueous solvent and an organic solvent, such as,but not limited to, the organic solvents as described above. Aqueoussolvents include, for example, water and buffered salts. In a preferredembodiment, the co-solvent includes about 70/30 ethanol/water by volume.In another preferred embodiment, the co-solvent includes from about60/40 to about 85/15 ethanol/water.

In a preferred embodiment the mixture includes a colloidal solution orcolloidal suspension. As used herein, the terms “colloidal solution” or“colloidal suspension” refer to a system intermediate between a truesolution and a suspension; the dispersed phase of the colloidal solutionor suspension have a particle size ranging from about 1 and 500nanometers.

In one embodiment of the invention, the mixture is prepared bysolubilizing a phospholipid in an organic solvent, such as, for exampleethanol, and a carboxylic acid or salt thereof in an aqueous solvent.The two phases are combined and a multivalent salt, such as, forexample, calcium chloride is added thereby forming a fine suspension orcolloidal solution.

The mixture can have a neutral, acidic or alkaline pH. Optionally, a pHbuffer can be added to the solvent or co-solvent or to the formedmixture. Preferably, the pH can range from about 3 to about 10. In oneembodiment, the mixture is a miscible mixture of organic and aqueousphases.

Suitable spray-drying techniques are described, for example, by K.Masters in “Spray Drying Handbook”, John Wiley & Sons, New York, 1984.Generally, during spray-drying, heat from a hot gas such as heated airor nitrogen is used to evaporate the solvent from droplets formed byatomizing a continuous liquid feed. Other spray-drying techniques arewell known to those skilled in the art. In a preferred embodiment, arotary atomizer is employed. An examples of suitable spray driers usingrotary atomization includes the Mobile Minor spray drier, manufacturedby Niro, Denmark. The hot gas can be, for example, air, nitrogen orargon.

Preferably, the particles of the invention are obtained by spray dryingusing an inlet temperature between about 100° C. and about 400° C. andan outlet temperature between about 50° C. and about 130° C.

Without being held to any mechanistic interpretation of the invention,it is believed that the formation of a colloidal solution facilitatesshell formation by (i) providing nucleation sites for the shell to formand (ii) slowing down the diffusion rates of the excipients in thedrying droplet. It is also believed that the presence of the carboxylatemoiety and calcium alters the phase behavior of the phospholipid insolution to create a colloidal aggregate phase that facilitates shellformation.

The spray dried particles can be fabricated with a rough surface textureto reduce particle agglomeration and improve flowability of the powder.The spray-dried particle can be fabricated with features which enhanceaerosolization via dry powder inhaler devices, and lead to lowerdeposition in the mouth, throat and inhaler device.

Particles including a medicament, for example one or more of the drugslisted above, are administered to the respiratory tract of a patient inneed of treatment, prophylaxis or diagnosis. Administration of particlesto the respiratory system can be by means such as known in the art. Forexample, particles are delivered from an inhalation device. In apreferred embodiment, particles are administered via a dry powderinhaler (DPI). Metered-dose-inhalers (MDI), nebulizers or instillationtechniques also can be employed.

Various suitable devices and methods of inhalation which can be used toadminister particles to a patient's respiratory tract are known in theart. For example, suitable inhalers are described in U.S. Pat. No.4,069,819, issued Aug. 5, 1976 to Valentini, et al., U.S. Pat. No.4,995,385 issued Feb. 26, 1991 to Valentini, et al., and U.S. Pat. No.5,997,848 issued Dec. 7, 1999 to Patton, et. al. Other examples include,but are not limited to, the Spinhaler® (Fisons, Loughborough, U.K.),Rotahaler® (Glaxo-Wellcome, Research Triangle Technology Park, N.C.),FlowCaps® (Hovione, Loures, Portugal), Inhalator® (Boehringer-Ingelheim,Germany), and the Aerolizer® (Novartis, Switzerland), the diskhaler(Glaxo-Wellcome, RTP, NC) and others, such as known to those skilled inthe art. Preferably, the particles are administered as a dry powder viaa dry powder inhaler.

The particles of the invention can be employed in compositions suitablefor drug delivery to the pulmonary system. For example, suchcompositions can include the particles and a pharmaceutically acceptablecarrier for administration to a patient, preferably for administrationvia inhalation. The particles can be co-delivered with larger carrierparticles, not including a therapeutic agent, the latter possessing massmedian diameters for example in the range between about 50 μm and about100 μm. The particles can be administered alone or in any appropriatepharmaceutically acceptable carrier, such as a liquid, for examplesaline, or a powder, for administration to the respiratory system.

The invention is also related to a method for drug delivery to thepulmonary system. The method comprises administering to the respiratorytract of a patient in need of treatment, prophylaxis or diagnosis aneffective amount of particles, such as described above, comprising atherapeutic, prophylactic or diagnostic agent. As used herein, the term“effective amount” means an amount required to achieve a desired effect,such as, for example, desired therapeutic response, or efficacy. Theactual effective amounts of drug can vary according to the specific drugor combination thereof being utilized, the particular compositionformulated, the mode of administration, and the age, weight, conditionof the patient, and severity of the symptoms or condition being treated.Dosages for a particular patient can be determined by one of ordinaryskill in the art using conventional considerations, (e.g. by means of anappropriate, conventional pharmacological protocol).

Aerosol dosage, formulations and delivery systems also may be selectedfor a particular therapeutic application, as described, for example, inGonda, I. “Aerosols for delivery of therapeutic and diagnostic agents tothe respiratory tract,” in Critical Reviews in Therapeutic Drug CarrierSystems, 6: 273-313, 1990; and in Moren, “Aerosol dosage forms andformulations,” in: Aerosols in Medicine, Principles, Diagnosis andTherapy, Moren, et al., Eds, Esevier, Amsterdam, 1985.

Preferably, particles administered to the respiratory tract travelthrough the upper airways (oropharynx and larynx), the lower airwayswhich include the trachea followed by bifurcations into the bronchi andbronchioli and through the terminal bronchioli which in turn divide intorespiratory bronchioli leading then to the ultimate respiratory zone,the alveoli or the deep lung. In a preferred embodiment of theinvention, most of the mass of particles deposits in the deep lung. Inanother embodiments of the invention, delivery is primarily to thecentral airways. Delivery to the upper airways can also be obtained.

In one embodiment of the invention, delivery to the pulmonary system ofparticles is in a single, breath-actuated step, as described in U.S.Patent Application, High Efficient Delivery of a Large Therapeutic MassAerosol, application Ser. No. 09/591,307, filed Jun. 9, 2000, which isincorporated herein by reference in its entirety. In another embodimentof the invention, at least 50% of the mass of the particles stored inthe inhaler receptacle is delivered to a subject's respiratory system ina single, breath-activated step. In a further embodiment, at least 5milligrams and preferably at least 10 milligrams of a medicament isdelivered by administering, in a single breath, to a subject'srespiratory tract particles enclosed in the receptacle. Amounts as highas 15, 20, 25, 30, 35, 40 and 50 milligrams can be delivered.

Porous or aerodynamically light particles, having a geometric size (ormean diameter) in the range of about 5 to about 30 μm, and tap densityless than about 0.4 g/cm³, such that they possess an aerodynamicdiameter of about 1 and about 3 μm, have been shown to display idealproperties for delivery to the deep lung. Larger aerodynamic diameters,ranging, for example, from about 3 to about 5 μm are preferred, however,for delivery to the central and upper airways. According to oneembodiment of the invention the particles have a tap density of lessthan about 0.4 g/cm³ and a mean diameter of between about 5 μm and about30 μm. According to another embodiment of the invention, the particleshave a mass density of less than about 0.4 g/cm³ and a mean diameter ofbetween about 5 μm and about 30 μm. In one embodiment of the invention,the particles have an aerodynamic diameter between about 1 μm and about5 μm. In another embodiment of the invention, the particles have anaerodynamic diameter between about 1 μm and about 3 μm microns. In stillanother embodiment of the invention, the particles have an aerodynamicdiameter between about 3 μm and about 5 μm.

In one embodiment of the invention, the particles are administered tothe respiratory system of a comatose, unconscious or anesthetizedpatient. In another embodiment, the particles are administered to therespiratory system of a nonhuman mammal, for example in veterinarymedicine or animal model experimental work. In a further embodiment ofthe invention, the particles are administered to sites other than thepulmonary system.

The present invention will be further understood by reference to thefollowing non-limiting examples.

Exemplifications

Some of the methods and materials employed in the following examples aredescribed in U.S. application Ser. No. 09/211,940, filed Dec. 15, 1998,in U.S. application Ser. No. 08/739,308, filed Oct. 29, 1996, now U.S.Pat. No. 5,874,064, in U.S. application Ser. No. 08/655,570, filed May24, 1996, in U.S. application Ser. No. 09/194,068, filed May 23, 1997,in PCT/US97/08895 application filed May 23, 1997, in U.S. applicationSer. No. 08/971,791, filed Nov. 17, 1997, in U.S. application Ser. No.08/784,421, filed Jan. 16, 1997, now U.S. Pat. No. 5,855,913 and in U.S.application Ser. No. 09/337,245, filed on Jun. 22, 1999, all of whichare incorporated herein by reference in their entirety.

Materials

Citric acid and calcium chloride were obtained from Spectrum Labs,Laguna Hills, Calif. DPPC was obtained from Avanti (Alabaster, Ala.).

Spray Drying

A Mobile Minor spray-drier from Niro (Denmark) was used. The gasemployed was dehumidified air. The gas temperature ranged from about 80to about 150° C. The atomizer speed ranged from about 15,000 to about50,000 RPM. The gas rate was 70 to 92 kg/hour and the liquid feed rateranged from about 50 to about 100 ml/minute.

Geometric Size Distribution Analysis

Size distributions were determined using a Coulter Multisizer II.Approximately 5-10 mg of powder was added to 50 mL isoton II solutionuntil the coincidence of particles was between 5 and 8%. Greater than500,000 particles were counted for each batch of spheres.

Aerodynamic Size Distribution Analysis

Aerodynamic size distribution was determined using anAerosizer/Aerodispenser (Amherst Process Instruments, Amherst, Mass.).Approximately 2 mg powder was introduced into the Aerodisperser and theAerodynamic size was determined by time of flight measurements.

Particle Morphology by Scanning Electron Microscopy (SEM)

Microsphere morphology was observed by scanning electron microscopy(SEM) using a Stereoscan 250 MK3 microscope from Cambridge Instruments(Cambridge, Mass.) at 15 kV. Microspheres were freeze-dried, mounted onmetal stubs with double-sided tape, and coated with gold prior toobservation.

Particle Density Analysis

Bulk density was estimated by tap density measurements, such as obtainedusing a Dual Platform Microprocessor Controlled Tap Density Tester(Vankel, N.C.) and confirmed by mercury intrusion analysis at PorousMaterials, Inc. (Ithaca, N.Y.).

EXAMPLE 1

300 milliliters of an aqueous solution containing 0.07% sodium citratebuffered to PH: 7.0 via addition of HCl was combined with 700milliliters of ethanol solution containing 0.1% DPPC. Four millilitersof a 2.5% aqueous CaCl₂ solution was added to the stirred mixture, atwhich point the colloidal solution was formed.

The mixture was spray dried. Inlet temperature was about 110 C., Feedrate about 60-70 m/min and atomizer spin rate 15000-20000 RPM. The tapdensity of the particles obtained ranged from 0.05 to 0.1 g/cm³. Yieldwas about 35-50%. The median geometric diameter of the resultingparticles was 10.7 microns and the median aerodynamic diameter was 2.2microns. SEM data of these particles indicated that they have a crumpledpaper like morphology.

EXAMPLE 2

The mixture was prepared and spray dried as described above. The aqueousphase (300 ml, 200 mg Na-Citrate, pH=7.0) and the ethanol phase (700 ml,700 mg DPPC) were mixed and stirred. CaCl₂ (25 mg/mg aqueous solution)was added dropwise. Amounts of calcium chloride used are shown and theproperties of the particles obtained are shown in Table 1.

TABLE 1 Amount of VMGD MMAD Est. Tap CaCl2 added Yield (%) (microns)(microns) Density  0 mg ~0 — — —  50 mg 15 6.62 3.35 0.26  75 mg 41 9.622.66 0.08 100 mg 36 9.72 2.39 0.06 125 mg 36 9.06 2.66 0.09

EXAMPLE 3

Particles containing albuterol sulfate were prepared in the followingmanner. A mixture including 66% DPPC, 20% sodium citrate, 10% calciumchloride and 4% albuterol sulfate was formed in a 70/30 (v/v)ethanol/water cosolvent system as described above and spray-dried. Themedian geometric diameter of the resulting particles was 9.2 microns andthe mass mean aerodynamic diameter was 2.5 microns.

While this invention has been particularly shown and described withreferences to preferred embodiments thereof, it will be understood bythose skilled in the art that various changes in form and details may bemade therein without departing from the scope of the inventionencompassed by the appended claims.

1. A method of preparing particles having a tap density less than about0.4 g/cm³ comprising: (a) forming a mixture including: a therapeutic,prophylactic or diagnostic agent, or any combination thereof, at least10% weight of a hydroxydicarboxylic acid and/or hydroxytricarboxylicacid or a salt thereof, a phospholipid, a salt comprising at least onemultivalent cation or anion and a solvent; and (b) spray-drying saidmixture to produce particles having a tap density less than about 0.4g/cm³ and an aerodynamic diameter between 1 and 5 microns.
 2. The methodof claim 1, wherein the particles have a median geometric diameter ofbetween about 5 microns and about 30 microns.
 3. The method of claim 1,wherein the particles have an aerodynamic diameter of between about 1and about 3 microns.
 4. The method of claim 1, wherein the particleshave an aerodynamic diameter of between about 3 and about 5 microns. 5.The method of claim 1, wherein the hydroxydicarboxylic acid and/orhydroxytricarboxylic acid is citric acid, or a salt thereof.
 6. Themethod of claim 1, wherein the phospholipid is endogenous to the lung.7. The method of claim 1, wherein the phospholipid is selected from thegroup consisting of phosphatidylcholines, phosphatidyletanolamines,phosphatidylglycerols, phosphatidylserines, phosphatidylinositols andcombinations thereof.
 8. The method of claim 1, wherein the phospholipidis present in the mixture in an amount of at least about 20% weight. 9.The method of claim 1, wherein the salt comprising at least onemultivalent cation or anion is a salt of an alkaline earth metal. 10.The method of claim 1, wherein the salt comprising at least onemultivalent cation or anion is a chloride.
 11. The method of claim 1,wherein the salt comprising at least one multivalent cation or anion iscalcium chloride.
 12. The method of claim 1, wherein the salt comprisingat least one multivalent cation or anion in present in the mixture in anamount of at least about 5% weight.
 13. The method of claim 1, whereinthe therapeutic, prophylactic or diagnostic agent is albuterol.
 14. Themethod of claim 1, wherein the therapeutic, prophylactic or diagnosticagent is present in the mixture in an amount between about less than 1%and about 40% weight.
 15. The method of claim 1, wherein the solventincludes an organic solvent.
 16. The method of claim 1, wherein thesolvent includes an aqueous organic co-solvent.
 17. The method of claim1, wherein the mixture is a colloidal solution.